Functional telomerase is required for functional iPS cells

Since the first report of induced pluripotent stem cells in 2006, the field of regenerative medicine has been buzzing about the potential for such cells to provide a source of cells that avoid the ethical minefield that plagues the use of embryonic stem cells.

The original paper demonstrated that simply over-expressing four “reprogramming” factors (4F) allowed reprogramming of differentiated mouse and human cells into iPS cells, but a thorough characterization of the resulting cells is still underway. iPS cells exhibit telomerase activity (as do ES cells), but whether this is sufficient to restore telomere length, or if telomeric chromatin acquires ES-like characteristics, remains unclear.

To address these issues, a team led by Maria Blasco generated iPS cells using either the standard 4 factors (4F), or omitting cMyc (3F), from wild-type and telomerase-deficient mice (both young and old), and investigated various aspects of telomere dynamics:

Telomere shortening is associated with organismal aging. iPS cells have been recently derived from old patients; however, it is not known whether telomere chromatin acquires the same characteristics as in ES cells. We show here that telomeres are elongated in iPS cells compared to the parental differentiated cells both when using four (Oct3/4, Sox2, Klf4, cMyc) or three (Oct3/4, Sox2, Klf4) reprogramming factors and both from young and aged individuals. We demonstrate genetically that, during reprogramming, telomere elongation is usually mediated by telomerase and that iPS telomeres acquire the epigenetic marks of ES cells, including a low density of trimethylated histones H3K9 and H4K20 and increased abundance of telomere transcripts. Finally, reprogramming efficiency of cells derived from increasing generations of telomerase-deficient mice shows a dramatic decrease in iPS cell efficiency, a defect that is restored by telomerase reintroduction. Together, these results highlight the importance of telomere biology for iPS cell generation and functionality.

The key findings were (1) in telomerase-competent cells, telomere lengthening occurs via telomerase extension (rather than via a recombination), and (2) telomeric chromatin acquires ES-like characteristics. Furthermore, cMyc (one of the original 4 transcription factors used to generate iPS cells ) is dispensible for telomerase activation in mouse iPS cells (telomerase activity was only marginally lower in the absence of cMyc).

The authors also derived iPS cells from telomerase-deficient G1 (first generation) mice. It became clear that while telomerase activity is not limiting for in vitro iPS cell proliferation when telomeres are long (as is the case in G1 mice), these cells were nonetheless severely impaired in their ability to generate viable mice. Furthermore, the efficiency of iPS cell generation from telomerase-deficient G2 and G3 mice dropped significantly, indicating that telomere shortening is a critical barrier to iPS cell generation. Consistent with this, the cells exhibited an increased number of signal-free ends and chromosome end-to-end fusions, both events that are common in cells with very short telomeres. Crucially, re-introduction of telomerase into G3 telomerase-deficient mice restored iPS cell production efficiency, despite the cell inheriting short telomeres from their G2 deficient parents.

Overall, these findings highlight the role that telomere/telomerase dynamics play in successful iPS cell generation and provide evidence that cells from older donors are suitable. The caveat is that telomerase activity is vital if iPS cells are to be generated from cells with short telomeres. (Obviously!)